HR+/HER2− breast cancer (BC) has a much lower pathological complete response (pCR) rate to neoadjuvant chemotherapy (NAC). Therefore, to better stratify the relapse risk for HR+/HER2− non-pCR populations, it is essential to accurate identification new prognostic markers.The study retrospectively analyzed 105 stage II–III patients who were diagnosed with HR+/HER2− BC and received NAC followed by breast and axilla surgery between 2013 and 2019 in Sun Yat-Sen University Cancer Center. The Miller–Payne (MP) grading system was used to evaluate pathological responses to NAC. The 70-gene signature was used to classify the prognosis signatures.p = 0.004, p = 0.021, and p = 0.001, respectively). Among 54 patients who underwent the 70-gene assays, 26 (48.1%) had a low-risk signature; 28 (51.9%) patients had a high-risk signature. Patients with poor response (MP grades 1–2) were more likely to with a high-risk 70-gene signature than those with good response (MP grades 4–5). The final analysis showed that DFS was longer in the low-risk group than in the high-risk group . DFS was longer in the good response (MP grades 3–4) group than in the poor response (MP grades 1–2) group . When stratified by MP grades combined with the 70-gene signature, subgroup analyses showed the good-response low-risk group with the best DFS, whereas the poor-response high-risk group showed the worst DFS (p = 0.048). Due to the short median follow-up time of 34.5 months (5.9–75.1 months), MP grades and the 70-gene signature did not show significant prognostic value for overall survival.Among the 105 patients, the study demonstrated that larger tumor size and lower progesterone receptor level at baseline and larger tumor size postoperative were statistically significantly associated with worse disease-free survival (DFS) (The study showed that analysis of MP grades combined with the 70-gene signature with residual NAC-resistant breast samples has a significant correlation with DFS. Hormone receptor-positive (HR+)/human epidermal growth factor receptor 2-negative (HER2−) breast cancer (BC) accounts for about 50–60% of all invasive breast carcinomas . HR+/HERThe 70-gene signature (MammaPrint) is a prognostic tool that classifies tumors into groups that are associated with a good prognosis or a poor prognosis based on the risk of distant metastases at 5 and at 10 years . The proIntegrating tumor size and nodal status with tumor grade and genomic signatures could provide accurate prognostic estimates for HR+/HER2− BC , 13. TheThe flowchart of the study is shown in The 70-gene expression profile was assessed from the formalin-fixed paraffin-embedded resection breast samples blinded to clinical and pathological data. The paraffin blocks were cut into tissue sections with a thickness of 3–5 µm to make white slices without cover glass and then dewaxed to obtain enough tumor RNA for genetic testing , 17. RNAThe pathological response to NAC was assessed by the MP grading system , compareGrade 1: No change or some alteration to individual malignant cells but no reduction in overall cellularity;Grade 2: A minor loss of tumor cells but overall cellularity still high; up to 30% loss;Grade 3: Between an estimated 30% and 90% reduction in tumor cells;Grade 4: A marked disappearance of tumor cells such that only small clusters or widely dispersed individual cells remain; more than 90% loss of tumor cells;in situ (DCIS) may be present.Grade 5: No malignant cells identifiable in sections from the site of the tumor, only vascular fibroelastotic stroma remains often containing macrophages. However, ductal carcinoma in situ residuals in breast tissue after surgery and complete disappearance of lymph node metastasis.pCR was defined as no histological evidence of malignancies or only Grades 1 and 2 are categorized as poor response, and Grades 3–5 are categorized as good response.The follow-up data including the dates of recurrence and death were collected through the outpatient service, telephone, and the way to the hospital review, until death or the date of the last follow-up . The median follow-up time was 37.5 months (2.4–92.3 months). DFS was calculated from the date of surgery to the date of disease relapse . OS was calculated from the date of surgery to the date of death or the latest follow-up.p < 0.05 were considered statistically significant.Analyses were performed using SPSS version 26. The differences in patients and tumor characteristics between the 70-gene high- and low-risk signature were tested using Fisher’s exact test. Logistic regression was applied to identify variables predictive of MP grades. The results of the model were expressed as odds ratio (OR) and relative 95% confidence interval (CI). Univariate and multivariate analyses were performed with the Cox proportional model to determine the effects of independent prognostic factors on DFS and OS. Calculating the exponential of the regression coefficients from the Cox model provided an estimate of the HR and the 95% CI. DFS and OS curves were calculated using the Kaplan–Meier method and compared using the log-rank test. Two-tailed We collected the information of 105 BC patients who received NAC before surgery 2. The mp = 0.042, OR 2.25, 95% CI 1.03–4.92; primary systemic therapy: p = 0.014, OR 4.48, 95% CI 1.35–14.84; clinical response: p = 0.010, OR 0.33, 95% CI 0.14–0.77), while it was unrelated to age, clinical tumor stage, clinical stage, PR, Ki-67, and molecular subtypes  to NAC in early-stage HR+/HER2− BC was significantly associated with N-stage 2–3, Anthracycline-taxane or clinical PR . The other factors were not related to good response. ER could not be analyzed for MP grades because most patients had tumors with high ER expression.Multivariate analysis was performed using baseline characteristics with The 70-gene signature was analyzed in 54 patients; the tumor characteristics are presented in Of the 105 early-stage HR+/HER2− cases, the magnitude of impact on different prognosis including DFS and OS differed depending on factors (p < 0.05); however, it was unrelated to histological grade, PR, KI67, molecular subtypes, nodal stage, pathological stage, MP grades, and MammaPrint signature . Postoperative clinicopathological factors were analyzed for survival Table 5.p = 0.037) (p = 0.018)  group than in the poor response (MP grades 1–2) group . However, a previous study showed that lower estrogen receptor level after 2 weeks of endocrine therapy is associated with poorer recurrence-free survival  score can be as a prognostic marker for long-term survival after NAC independent of the molecular subtype . The prop = 0.037, p = 0.018, respectively). Stratifying by MP grades and the 70-gene signature also showed marked prognostic value (p = 0.048). The poor-response high-risk group showed the worst DFS, whereas the good-response low-risk group showed the best DFS. All patients received tamoxifen or aromatase inhibitors combined with ovarian function suppression adjuvant endocrine therapy. The study showed that active endocrine intervention for the poor-response high-risk group was not enough; the addition of chemotherapy or intensive endocrine therapy  in the adjuvant setting may prolong DFS and improve prognosis for these patients. Due to the short follow-up time, MP grades and the 70-gene signature did not show significant prognostic value for OS. A previous study used tumor samples prior to NAC for the 70-gene, which signature showed good prognostic value and predictive chemotherapeutic sensitivity  were more likely to have a high-risk 70-gene signature than those with good response (MP Grades 4–5) Figure 1sitivity . HoweverThere were several limitations that should be considered while interpreting the study findings. One critical limitation was the small sample size that restricted our analysis for the subgroup based on MP grades and the 70-gene signature. Another limitation was that a longer follow-up will be required to confirm that MP grades and the 70-gene signature were associated with long-term survival. Different NAC regimens and cycles were another issue. In the present study, most patients received different cycles of anthracycline-based or anthracycline-taxane NAC, which may affect a reduction in tumor load of the primary tumor. Despite these limitations, our study has several strengths. To our knowledge, this is the first study that provides an analysis of MP grades combined with the 70-gene signature associated with prognostic in HR+/HER2− non-pCR populations. Integration of MP grades and the 70-gene signature to determine the appropriate options of adjuvant treatment may prolong DFS and improve prognosis. Further studies are warranted to confirm the clinical utility of MP grades combined with the 70-gene signature in early-stage HR+/HER2− breast cancer.In conclusion, this study showed that analysis of MP grades combined with the 70-gene signature with residual neoadjuvant chemotherapy-resistant breast samples has a significant correlation with DFS. In particular, subgroup analyses based on MP grades and the 70-gene signature also showed markedly prognostic value.The original contributions presented in the study are included in the article/Conception and design of the study: FX and SW. Acquisition of clinical data: LW, QL, KJ, RH, KL, PZ, and DZ. Miller–Payne grades pathological evaluation: RL. Analysis and interpretation of the data: LW and QL. Manuscript drafting and revision: LW, QL, FX, and SW. All authors contributed to the article and approved the submitted version.The work was funded by the Sun Yat-sen University Clinical Research 5010 Program (2017010).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


The energy-efficient deep ultraviolet (DUV) optoelectronic devices suffer from critical issues associated with the poor quality and large strain of nitride material system caused by the inherent mismatch of heteroepitaxy. In this work, we have prepared the strain-free AlN film with low dislocation density (DD) by graphene (Gr)-driving strain-pre-store engineering and a unique mechanism of strain-relaxation in quasi-van der Waals (QvdW) epitaxy is presented. The DD in AlN epilayer with Gr exhibits an anomalous sawtooth-like evolution during the whole epitaxy process. Gr can help to enable the annihilation of the dislocations originated from the interface between AlN and Gr/sapphire by impelling a lateral two-dimensional growth mode. Remarkably, it can induce AlN epilayer to pre-store sufficient tensile strain during the early growth stage and thus compensate the compressive strain caused by hetero-mismatch. Therefore, the low-strain state of the DUV light-emitting diode (DUV-LED) epitaxial structure is realized on the strain-free AlN template with Gr. Furthermore, the DUV-LED with Gr demonstrate 2.1 times enhancement of light output power and a better stability of luminous wavelength compared to that on bare sapphire. An in-depth understanding of this work reveals diverse beneficial impacts of Gr on nitride growth and provides a novel strategy of relaxing the vital requirements of hetero-mismatch in conventional heteroepitaxy. This work successfully achieves a strain-free AlN film with low dislocation density for DUV-LED through graphene-driving strain-pre-store engineering and present the unique mechanism of strain-relaxation in QvdW epitaxy. Therefore, achieving a high-quality epitaxy of AlN films is of particular importance to ensure the excellent performance of DUV photoelectric devices13. Currently, due to the lack of large-size and low-price homogenous substrates, the optimal choice to grow AlN films is usually to perform heteroepitaxial growth on sapphire17. Unfortunately, due to the inherent lattice and thermal expansion coefficient (TEC) mismatches between AlN and sapphire substrate19, even the well-known two-step epitaxy method or epitaxial lateral overgrowth (ELO) technology still inevitably introduces a variety of crystal defects into AlN epilayer22. In particular, the large residual strain in the AlN film leads to the nonuniformity of the Al distribution in the upper AlGaN layer accompanied by wafer bending, which severely limits the device performance24. The way to release large residual strain of the film is to mitigate the strong interactions between the substrate and the epilayer, or to introduce “negative strain” to neutralize the original residual strain26. Therefore, a more feasible strategy is straightaway required to make a qualitative leap to realize high-quality growth of heteroepitaxial AlN films and to meet the application requirements of DUV optoelectronic devices.The direct band gap of AlN-based materials makes them suitable for fabricating optoelectronic devices30. The widely studied 2D material, graphene (Gr) has been incorporated as a buffer layer for the epitaxial growth of nitrides to effectively alleviate the lattice and TEC mismatches between the epilayer and the substrate33. Besides, it is worth mentioning that the metal adatoms of nitrides have a tiny migration barrier on the Gr surface, which can undoubtedly promote the 2D growth trend of nitride films35. Kim et al. demonstrated direct vdW epitaxy of high-quality single-crystalline GaN film on epitaxial Gr with low defects, and realized the epilayer transfer from Gr/SiC substrates36. Chung et al. achieved the growth of high-quality GaN films utilizing Gr/ZnO nanowalls as an interlayer and fabricated GaN-based blue LED that can be transferred to foreign substrates37. However, the previously reports on the epitaxial nitride film on Gr usually stated that the stress relaxation of epitaxial system is realized through the weak interaction between Gr itself and epilayers, but there is no detailed discussion or rigorous verification of this statement38. Until 2020, Bae et al. reported the discovery of a novel pathway of relaxing misfit strain in heteroepitaxial films via interface displacement on Gr-coated substrates25. The aforementioned material systems are those with relatively small lattice mismatch (InGaP on GaAs with 0.74% mismatch and GaP on GaAs with 3.7% mismatch) and the Gr undergoes a complicated transfer process instead of directly growing on the heterogeneous substrate. However, the potential of Gr film as an interlayer to release the strain in a large-mismatched epitaxial material system  is still ambiguous and needs further development. Meanwhile, Dou et al. observed the chemical bond formation at the interface between the directly grown Gr and sapphire by aberration-corrected transmission electron microscopy (TEM) and found the strong interaction between Gr and sapphire39, which will inevitably change the interface displacement of Gr during QvdW growth. Therefore, there must be a novel QvdW epitaxial mechanism to explore for nitrides on the directly grown Gr-covered substrate. Unfortunately, there have been few reports to reveal the QvdW epitaxy mechanism of AlN films, which is essential to precisely manipulate the epitaxial quality of AlN films and further elevate the performance of DUV optoelectronic devices.Over recent years, an emerging method named quasi-van der Waals (QvdW) epitaxy or remote epitaxy based on two-dimensional (2D) material has been proposed for high-quality heteroepitaxial growth of group-III nitrides25. According to the results obtained by X-ray diffraction (XRD) and TEM, the DD of AlN with Gr exhibits an anomalous sawtooth-like evolution during the QvdW epitaxy process and the values are consistently lower than that without Gr. More importantly, combining Raman analysis and first-principles calculations, it is revealed that plasma-treated Gr with a high density of small-size AlN nucleation islands will pre-store sufficient tensile strain during the coalescence process. The tensile strain can compensate the compressive strain caused by lattice and TEC mismatches during heteroepitaxy, thus bringing out a strain-free AlN film. The reciprocal space mapping (RSM) of the as-fabricated DUV light-emitting diode (DUV-LED) reveals only a weak compressive strain in the n-AlGaN layer, enabling the high-quality crystalline state of the upper LED structure. The as-fabricated 283 nm DUV-LED with Gr demonstrates 2.1 times higher light output power (LOP) compared to its counterpart on bare sapphire and better stability of luminous wavelength under the changing injection current. This work reveals the internal mechanism of QvdW growth of AlN on large-mismatched substrate and undoubtedly sheds light on the further promotion of nitride-based device manufacturing.Herein, we successfully achieve a strain-free AlN film with low dislocation density (DD) through Gr-driving strain-pre-store engineering and present the unique mechanism of strain-relaxation in QvdW epitaxy, different from the previous “interface displacement” argument2 and 30 sccm CH440. Figure 2/Si substrate exhibits the same color contrast that indicates its good uniformity  sapphire via a catalyst-free atmosphere chemical vapor deposition (CVD) method at 1050 °C for 3 h, under a gas mixture of Ar, Hity Fig. . And theity Fig.  as well ity Fig.  of the gent Fig. . The ave sapphire2 plasma treatment, and then the epitaxy of the AlN film is performed. The nucleation of AlN mainly occurs at the N defect sites through Al–N bond as the fulcrum, and then AlN clusters eventually rapidly expand into the film in the form of vdW epitaxy driven by Gr6. For the growth on bare sapphire, the AlN epilayer is initiated with the thin nucleation at high temperature . It is attributed to the adoption of N2 plasma treatment prior to growth, which can greatly facilitate AlN nucleation by introducing pyrrolic-nitrogen defects into Gr6. Furthermore, since Al atoms have an extremely low migration barrier (less than 0.1 eV) on Gr34, the nucleated AlN rapidly coalesces to fully cover the Gr-buffered sapphire  of the X-ray rocking curve (XRC) of AlN filmO3 Fig. S. Meanwhi2 (high) phonon mode in the Raman spectrum, as depicted in Fig. 45. In the initial growth stage, regardless of the presence of Gr, the position of E2 (high) peak blue-shifts continuously. For the AlN epilayer about 700 nm on sapphire, the position of E2 (high) peak suddenly red-shifts. In contrast, the tensile strain accumulation of the AlN epilayer on Gr is more intense in the initial stage, so the turning point of E2 (high) peak of the AlN epilayer on Gr appears earlier at 600-nm thickness, followed by a similar sharp increase at 700 nm. When the position of E2 (high) peak shifts to a higher wavenumber for 700-nm-thick AlN epilayer, the strain state of the epilayer has changed from tensile to compressive strain. Therefore, it is reasonable to speculate that the regeneration of the dislocations at a thickness of about 600–700 nm observed in the TEM originates from the release of the accumulated tensile strain in the epilayer. The residual compressive strain should be attributed to the TEC mismatch during the cooling process. More importantly, from a macro perspective, the wavenumber of the E2 peak of AlN epilayer with Gr is always smaller than that of AlN on bare sapphire under the same thickness, suggesting that the presence of Gr provides an additional source of tensile strain in the epitaxy system. Consequently, the strain-free AlN film based on Gr is eventually obtained, while AlN film on bare sapphire demonstrates the residual compressive strain. In addition, RSM by high-resolution XRD is ideally suited to detailed structural characterization of layered nitride structure. As shown in Fig. S46.In order to understand the stress relaxation of AlN films grown on sapphire with and without Gr during the heteroepitaxy, we explore the strain state of AlN epilayer with different growth thicknesses by characterizing the stress-sensitive E47, it is determined that the positive lattice mismatch of +13.3% and the negative TEC mismatch of −44% inevitably exist between AlN and c-surface sapphire, which enforces the AlN epilayer grown on sapphire to exhibit a compressive strain. On the other hand, there exists a source of tensile stress during the coalescence process. As the atoms on the crystalline planes from different nuclear islands start to form chemical bonds with each other, the contact and coalescence of adjacent AlN clusters will inevitably induce the tensile strain. And the magnitude of the local tensile strain introduced in the coalescence process depends on the size of the nuclear islands that coalesce with each other48. Hence, in order to investigate the characteristics of the tensile strain between adjacent AlN clusters developed during the coalescence process, we performed first-principles calculations based on density functional theory (DFT) to study the structural relaxation of two surfaces at different separations. The most stable side facet of the AlN clusters is the  =1.94 Å, 0.3 Å longer than the bond length in the bulk.We first consider the bonds parallel to the coalescent surfaces. At a larger gap, these surface bonds are always shorter than those in bulk due to surface contraction, and slightly elongate when the gap decreases, as shown in Fig. d//,c(Al) and d//,c(N)) are longer than those of infinite surface by about 0.04 Å at the critical gap of 3.6 Å. This indicates that the tensile strain of small AlN clusters is larger than that of large clusters during the coalescence. The larger elongation of the corner bonds on nanowires is due to a higher degree of freedom of the corner atoms. Compared to the atoms on the infinite surface, the corner atoms of a nanowire can move outwards by a larger distance.In nanowires, the surface bonds at the edge and near the corner exhibit different variations as the gap decreases. The elongation of the surface-parallel bonds on the edge of the nanowire is similar to those on the infinite surface Fig. S. Howeverd⊥,e(Al) and d⊥,e(N)) are elongated by a larger amount than those on the infinite surface. The perpendicular bonds on the corner of the nanowire are essentially the surface bonds on the neighboring facets and thus have a smaller elongation , which becomes more and more stronger as the epilayer grows thicker. To a certain extent, the tensile strain dominates the AlN epilayer even after cooling down to RT . However, although the state of tensile strain is maintained until the AlN film is completely formed at high temperature , the AlN epilayer grown on sapphire eventually exhibits compressive stress due to thermal mismatch during the cooling process as shown in Fig. 24.Based on the characterization analysis and DFT calculations mentioned above, a schematic growth model is proposed to elucidate the internal mechanism of Gr to affect the crystallinity and strain state of the AlN epilayer. As mentioned above, negative thermal mismatch will introduce compressive strain into the AlN epilayer. Therefore, in order to interpret the results of the Raman spectrum measured at room temperature (RT) more straightforwardly, the strain state of the epilayer at each stage from the cooling down to the growth interruption is also depicted. As shown in Fig. 5a RT Fig. . After ture Fig. 0, the Al2 plasma-treated Gr promotes the formation of high-density and small-size nucleation islands of AlN. On one word, more coalescence interfaces between islands will undoubtedly induce larger tensile strain compared to its counterpart on bare sapphire. On the other word, according to first-principles calculation results, smaller-size AlN nucleation islands can also trigger more intense tensile strain during the coalescence process. As a result, even after partial strain release by the formation of dislocations, the AlN epilayer still suffers from a large tensile strain  with AlN/Gr shows a tiny deviation from the intrinsic AlGaN  with the Al composition of 0.55, suggesting that there is only a weak biaxial compressive strain in n-AlGaN layer49. However, the n-Al0.55Ga0.45N layer  contained in the DUV-LED on bare sapphire shows the existence of greater compressive strain  layers, thus ensuring its high internal quantum efficiency (IQE). At 20 mA, the LOP of the DUV-LED with Gr reaches 1.26 mW, which is 2.1 times higher than that without Gr (0.60 mW), confirming the high quality of the DUV-LED device due to the Gr buffer layer. According to the calculation, when the applied current is 20 mA, the corresponding external quantum efficiency (EQE) and wall-plug efficiency (WPE) of DUV-LED with Gr buffer layer are 1.44% and 1.02%, respectively, which are both higher than the EQE (0.68%) and WPE (0.5%) of LED without Gr. Moreover, the current-voltage curve of the DUV-LED with Gr exhibits a good rectifying behavior with a turn-on voltage of ~4.6 V In summary, through both experimental and theoretical analysis of DFT calculation, this work reveals the unique mechanism of strain-relaxation of AlN epilayer in QvdW epitaxy guided by Gr-driving strain-pre-store engineering. Furthermore, the epitaxial LED structure on the strain-free AlN also exhibits a low residual strain, and the DUV-LED based on strain-free AlN/Gr shows more excellent and more stable optoelectronic properties than that on bare sapphire, demonstrating the great application of the strain-free AlN film to high-performance DUV-LED. These valuable perceptions about the inherent potence of Gr related to the growth of nitride film undoubtedly provide fulfilling enlightenment for developing the practical application of Gr in the cutting-edge manufacturing of nitride-based optoelectronic devices.2. Then 30 sccm CH4 is introduced into the reaction chamber as a carbon source for the growth of Gr on the sapphire substrate for about 3–5 h.Typically, a two-inch sapphire substrate is cleaned with deionized water, ethanol, and acetone and then loaded into a three-zone high-temperature furnace. The furnace is heated to 1050 °C and stabilized for about 10 min under 500 sccm Ar and 300 sccm H2 plasma treatment  under optimized plasma treatment conditions (200 Pa pressure with 300 sccm air flow and 50 w power for 30 s) before loading into MOCVD chamber. And the type of plasma treatment is reactive ion etching (RIE) which is capacitively coupled radio frequency (RF) plasma. Table 3 flow of 500 sccm and a trimethylaluminum (TMAl) flow of 70 sccm on sapphire with and without the Gr interlayer in a single experiment. And the V/III ratio of the subsequent growth process is 270. It is a one-step process without using LT AlN buffer layer.The Gr/sapphire is exposed to N0.6Ga0.4N superlattice (SL), n-Al0.55Ga0.45N layer, five-period Al0.4Ga0.6N/Al0.5Ga0.5N MQWs, and p-type layers (layer of Mg-doped p-Al0.65Ga0.35N electron blocking layer (EBL), p-Al0.5Ga0.5N cladding layer and p-GaN contact layer). Trimethylgallium (TMGa) is used as a Ga precursor. Silane (SiH4) and bis (cyclopentadienyl) magnesium (Cp2Mg) are used for n-type and p-type doping, respectively. A 20-period AlN (60 nm)/Al0.6Ga0.4N (15 nm) SL is first deposited at 1130 °C, with the periodic flow change of TMAl to adjust the deposition component while the TMGa flow is kept at 32 sccm. Then temperature is reduced to 1002 °C, and 20 sccm silicane hydrogen mixture flow  is introduced for the growth of the 1.8 μm n-Al0.55Ga0.45N layer. The five-period Al0.5Ga0.5N/Al0.4Ga0.6N MQWs structure is further grown with a 3 nm quantum well and a 12 nm quantum barrier by switching the TMAl from 24 to 14 sccm and TMGa from 8 to 7 sccm for each period. A 50 nm thick layer of Mg-doped p-Al0.65Ga0.35N EBL, a p-Al0.5Ga0.5N (30 nm) cladding layer, and a 100 nm thick p-GaN contact layer are subsequently deposited. After the growth, the p-type layers are annealed in the reactor at 800 °C in N2 atmosphere for 20 min to activate the Mg acceptors. In addition, the doping concentration of Si in the n-Al0.55Ga0.45N layer is 3 × 1018 cm−3 and the doping concentration of Mg in p-GaN is 1 × 1018 cm−3. Moreover, the corresponding estimated carrier concentrations of n-type layer and p-type layer are 3 × 1018 and 1 × 1016 cm−3, respectively.The MOCVD system used in the epitaxial growth process of DUV-LEDs is a home-made vertical system. The AlGaN-based DUV-LED structure is grown on the AlN/Gr/sapphire template, including a 20-period AlN/Al3/Cl2/Ar mixed gas, and the etching depth was about 600 nm. 2. Fabrication of N electrode and p electrode: a combination method of negative-photoresist photolithography and metal stripping is adopted. By means of electron beam evaporation, the Ti/Al/Ti/Au metal stack is deposited on the exposed n-AlGaN as the n-type contact, while the Ni/Au stack is used as the p-type contact. 3. Passivation and isolation: SiO2 as isolation layer is deposited by PECVD to reduce leakage and avoid short circuit. Finally, the DUV-LED chips are flip-chip bonded onto ceramic submounts coated with gold for light output testing.DUV-LED devices with a die size of 0.5 mm × 0.5 mm are fabricated following the standard LED processes. It includes the following steps: 1. Mesa etching: firstly, the positive-photoresist is used for photolithography of mesa pattern. Then, the photolithography pattern is further ICP etched with BClThe cross-sectional TEM specimen is prepared by the focused ion beam system (ThermoFisher Helios G4 UX). The HRTEM images, DF images, and SAED patterns are performed on FEI Tecnai F20 TEM operated at 200 kV. And the HAADF images and EDS mapping are acquired at a spherical aberration-corrected FEI electron microscope (Titan Cubed Themis G2 300) operated at 300 kV. The camera length in HAADF mode is set as 145 mm. The convergence semi-angle of HAADF is 30 mrad and the collection semi-angle of HAADF is 39–200 mrad.52. We adopted the projector-augmented wave potentials and the generalized-gradient approximation of Perdew, Burke, and Ernzerhof for the exchange-correlation functional54. The coalescence between small clusters is modeled by two AlN nanowires with Supplementary Information For Graphene-driving strain engineering to enable strain-free epitaxy of AlN film for deep ultraviolet light-emitting diode


The composite material (PANI-GO-APTES) was synthesized by chemical method and was characterized with SEM, XPS, XEDS, XRD, TGA, FTIR, EIS and CV. The result of characterization indicates the successful synthesis of the intended material. The PANI-GO-APTES was successfully applied for electrochemical detection of Pb2+ using cyclic voltammetry and linear sweep voltammetry method. The limit of detection of Pb2+ was 0.0053 µM in the linear range of 0.01 µM to 0.4 µM. The current response produced during the electrochemical reduction of Pb2+ catalyzed by PANI-GO-APTES was also very repeatable, reproducible and rapid. The application of PANI-GO-APTES-modified GCE in real sample analysis was also established. Therefore, PANI-GO-APTES is presented as a potential Pb2+ sensor for environmental and human health safety.Lead is a potentially toxic element (PTE) that has several adverse medical effects in humans. Its presence in the environment became prominent due to anthropogenic activities. The current study explores the use of newly developed composite materials (organic–inorganic hybrid) based on PANI-GO-APTES for electrochemical detection of Pb Hybrid organic–inorganic (HOI) composites were discovered several decades back. However, they did not attract the interest of the scientific community until the 1990s when Toyota recorded a massive increase in the strength of the materials used for its automobiles by incorporation of mica to nylon to yield novel HOI material . HOI matThe unrivaled excellent properties of graphene and graphene oxide such as large surface area, high electrical conductivity, high potential adsorption power and good catalytic properties make them efficient materials for several applications, including supercapacitors, energy storage devices and electrochemical sensing ,14,15,16Lead is primarily introduced into the environment by industrial activities such as industrial operations at mining sites. Other sources include lead battery manufacturers, lead-based pigment manufacturers, automobile industries, decayed water-carrying metal pipes and contaminated consumer products . ExposurExposure to lead has been implicated in an average of 143,000 deaths annually . Because2+ detection in environmental samples. Include the atomic absorption spectrophotometry method and the inductively coupled plasma method. The high cost of instrumentation and cumbersome sample preparation limit the use of the aforementioned methods. Other methods including UV-Vis spectroscopy and fluorescence spectroscopy have been used [2+ and Cu2+ had an influence on the current response of the working electrode towards lead (II) ion especially at high concentration. Anambiga and his coresearchers worked on lead (II) ion detection using a glutathione–silver-modified screen-printed electrode that employed the electrochemical method [The conventional methods for toxic metals, including Pbeen used ,32,33,34een used . Severaleen used , lead  through the chemical method. The obtained GO-APTES was then integrated into the PANI matrix precursor to form an organic–inorganic hybrid. The as-prepared PANI-GO-APTES was then applied as a lead electrochemical sensor at trace concentration. This study, as far as we know, reports for the first time the synthesis of the composite under study (PANI-GO-APTES) and its use for the detection of lead ions in aqueous solution.−1). The reagents used for this study, namely graphene oxide, aniline monomer, potassium persulfate (PPS), 3-aminopropyltriethoxysilane , hydrochloric acid, ethanol (EtOH), acetate buffer, lead sulfate, nickel chloride, copper sulfate, cadmium sulfate, manganese sulfate, sodium nitrite, sodium hydrogen phosphate and sodium hydroxide, were supplied by Sigma-Aldrich  and Alfa Aesar . The reagents were used for the study as received. The reagents were prepared using deionized water , energy-dispersive X-ray spectroscopy system , Fourier transform infrared spectrometer , Sonicator , thermogravimetric analyzer , X-ray diffractometer , X-ray photoelectron spectroscope  and 4-probe conductivity meter  electrochemical workstation .In this study, wet chemical polymerization method was used for PANI synthesis. The typical procedure involved the preparation of three different PANI samples by mixing aniline (varying concentrations) and oxidizing agent (PPS) in a ratio of 1:3 under continuous stirring for two hours until the color of the suspension changes to green.v/v) under sonication for 1 h in standard flask. After a homogenous suspension of GO has been achieved, 2 mL of APTES was added to the GO dispersion and allowed to interact. The mixture GO-APTES was then refluxed for 24 h maintaining 70 °C temperature.Two hundred milligrams of GO was dispersed in ethanolic solution  in the 2-theta angle range of 10–80° and scan rate of 0.05°/min. The elemental composition was investigated using XPS and EDX. For the XPS analysis, Al K monochromatic X-radiation source was used with a beam spot of 300 µm. EDX was fitted with SEM and simultaneously recorded with SEM imaging.−1 using KBr pellets.FTIR analysis was conducted to reveal the functionalities of the as-prepared PANI-GO-APTES. The spectra were collected at the IR wavenumber of 400–4000 cmTGA analysis was carried out to reveal information about the thermal stability of the synthesized sample. It was carried out in a nitrogen atmosphere in the heating range of 50–900 °C and heating rate of 20 °C/min and a gas flow of 20 mL/min. Electrical conductivity of the as-prepared PANI-GO-APTES was measured using a 4-probe conductivity meter. The as-prepared PANI-GO-APTES was made into pellets using a presser by applying a pressure of 5 kN with hydraulic pressure of 20 min. The produced pellets were then used for conductivity measurements.Electrochemical characterization and measurements were carried out using Autolab potentiostat. The typical potentiostat contains three electrodes: working, reference and counter electrodes. The working electrode was either a bare glassy carbon electrode (GCE) or PANI-GO-APTES-modified GCE at any point in time in this study. The reference electrode was a silver/silver chloride electrode system, and the counter electrode was a platinum wire. Detection of lead ions was carried out by electrochemical means. Specifically, cyclic voltammetry and linear sweep voltammetry methods were used. For the CV study, the potential window ranged from −1.2 to 1.0 V, using a scan rate of 75 mV/s and amplitude of 0.005 V. The linear sweep was used for calibration study and was carried out maintaining the potential window range of −0.8 to 0 V at a scan rate of 75 mV/s.2+ was based on PANI-GO-APTES-modified GCE. To start with, the bare electrode was thoroughly washed with distilled water and ethanol and electrochemically cleaned in 0.25 M H2SO4 (by CV). To the clean bare GCE, 10 µg of PANI-GO-APTES powdery material was dispersed in 10 µL Nafion solution. The dispersed PANI-GO-APTES/Nafion solution was then cast on the GCE and was allowed to anneal in the oven for about 10 min. The PANI-GO-APTES-GCE was then stored and used for further studies.The designed electrochemical sensor probe for PbThe obtained SEM image results are presented in The XEDS spectrum obtained for PANI-GO-APTES is shown in In another effort, XPS was carried out to determine the binding energy of the material under study (PANI-GO-APTES). XPS is a powerful tool for surface chemical investigation. The full XPS spectrum recorded for PANI-GO-APTES a indicatCarbon is deconvoluted into four peaks, namely C-O-C/C-O-Si, C-C/C=C, C-N and C=O corresponding to 286.89, 284.6, 285.47 and 288.5 eV, respectively , as presThe nitrogen spectra are deconvoluted into two peaks, namely C-N and H-N corresponding to the peaks at 401.9 and 403 eV, respectively d. MoreovThe structural investigation of the synthesized PANI-GO-APTES was studied with XRD. The obtained XRD spectrum is presented in −1 corresponds to N-H vibration of PANI and APTES. In addition, the peaks at 1638 and 1370 cm−1 could be attributed to quinoid and benzenoid vibrations, which implies effective interaction between GO-APTES and PANI. The observed peak at 1630 cm−1 may be due to N-H stretching vibration in PANI and APTES. Interaction between APTES and GO could bring about Si-O-Si stretching vibration, resulting in the peak at around 1097 cm−1. This peak could imply successful anchoring of the Si of APTES on the GO surface. The C-H stretching of APTES, PANI and GO could be associated with the peaks at 2928 and 2360 cm−1. The obtained results are consistent with the literature [terature ,44,45.The obtained TGA spectrum for the as-prepared PANI-GO-APTES is presented in 2+ to zerovalent Pb. The circuit diagram for the Nyquist plot and the values for the circuit parameters are presented in The EIS spectrum recorded for PANI-GO-APTES in 1 mM ferricyanide is presented in 7 (W/S) is the correction factor used for the case of a nonconducting bottom surface, which is a function of W, S the probe spacing (cm) and the thickness of the sample under test (cm).−1) and V is the voltage (V).The electrical resistivity of the as-prepared sample was measured using the four-probe method. The obtained current–voltage data were used to measure electrical resistivity using the following equations (Equations (1)–(4)) :(1)ρ=ρ˳∕The obtained results are summarized in 2+ with a distinguished high reduction current at −0.4 V. Other metal ions such as Ni2+, Cd2+, Zn2+ and Cu2+ had very little or no reduction current at this potential. Therefore, it was concluded that PANI-GO-APTES is sensitive and selective towards Pb2+. Hence, further electrochemical detection studies were carried out for Pb2+.2+. From the figure, it can be observed that PANI-GO-APTES displayed a distinguished high reduction current at −0.4 V. As compared to GO-APTES composite, the reduction current was higher, and this implies that incorporation of PANI into GO-APTES improved the electroreduction/catalytic property of GO-APTES. As reported elsewhere, PANI has excellent catalytic properties, which makes its use in electrochemical sensing very promising [2+. Equally, PANI-GO-APTES did not present reduction current at −0.4 V in the absence of Pb2+ (acetate buffer only). From this control study, it is established that the observed reduction current at −0.4 V was due to PANI-GO-APTES.romising ,48,49. O2+. The optimum reduction current was recorded with acetate buffer of pH 5.6. This suggests that electroreduction of Pb2+ was more favorable in an acidic buffer medium. On the other hand, in the alkaline medium (result not included), a viscous white precipitate was immediately formed in the electrochemical cell upon spiking with Pb2+. This resulted in no or very low current response. Therefore, the pH of the supporting electrolyte was maintained in the range of 7.0 to 3.6. From the result, the lowest reduction current was observed with pH 5.6. Therefore, the electrochemical studies for Pb2+ detection were conducted using the optimum pH medium (pH 5.6).Scan rate gives vital information on the electrochemical behavior of an analyte on the electrode surface . For insFor confirmation of the nature of the electrochemical reaction occurring on the electrode surface, the plot of the logarithm of reduction current response against the logarithm of scan rate was plotted f. The eqAs given in Equation (1), the slope of the plot is 0.46; slopes of <0.5 indicate a fully diffusion-controlled reaction, while those >0.5 indicate adsorption-controlled reactions . For the2+ is presented in 2+, no reduction peak was observed at −0.4 V. However, upon addition of 0.01 µM Pb2+, a reduction peak was observed. The peak increased linearly up to 0.4 µM. The plot of varying Pb2+ concentration against the reduction current is presented in The recorded current response of PANI-GO-APTES towards varying concentrations of Pb2+ on the surface of PANI-GO-APTES might be due to electroreduction of Pb2+ to zerovalent Pb catalyzed by PANI-GO-APTES substrate. The reaction is illustrated in Equation (8):The obtained correlation (0.99) suggests a good linear response in the linear range of 0.01 to 0.4 µM. The mechanism of detection of Pb−1 respectively. Moreover, the linear dynamic range and full dynamic range for the calibration plot were 0.01 to 0.4 µM and 0.01 to 1.0 µM, respectively.The merit parameters of interest for this study are limit of detection (LOD), limit of quantification (LOQ), sensitivity, linear dynamic range (LDR) and full dynamic range (FDR). The LOD, LOQ and sensitivity were calculated using the formula given in 2+ detection was examined by repeatability, reproducibility, interference studies and response time. 2+ concentration. The RSD of the recorded reduction peak current was 1.5%. The stability of PANI-GO-APTES-modified GCE for Pb2+, Cu2+, Mn2+, Zn2+, Cd2+ and other electroactive species such as nitrite and phosphate ions was investigated, and the obtained result is given in 2+ is very repeatable, reproducible and interference-buffered.The probable effect of likely interferents such as Ni2+ detection is presented in 2+ as it has a short response time desirable for in situ detector/sensors.In addition, the response time of PANI-GO-APTES in Pb2+ was employed. The percentage recovery of the spiked Pb2+ was then estimated using the formula stated in 2+ ranged from 86.7 to 106.7%. The obtained result suggests that PANI-GO-APTES can be used for the real detection of Pb2+.The real samples used for this study were tap water, well water and bottled water. A standard addition method of spiking the electrochemical cell with a known amount of Pb2+ in the environment due to its medical importance. Some of the pertinent work related to Pb2+ detection is displayed in As indicated in 2+ selectively using cyclic and linear sweep voltammetry methods. The minimum detectable concentration of lead was 0.0053 µM in the linear range of 0.01 to 0.4 µM. The sensitivity recorded was 165.71 µA µM−1cm−1, and the response time was less than 20 s. In the presence of likely interferents such as metal ions and electroactive species, PANI-GO-APTES current response towards Pb2+ was stable. The proposed method was also successfully applied for Pb2+ detection in real environmental water samples  with good percentage recovery. Therefore, this study established the application of PANI-GO-APTES-modified GCE for electrochemical detection of lead with good performance. The outcome of this research is a step in achieving a cheap, sensitive and accurate method for detection of lead in the environment to improve human health and protect the environment.In this study, novel PANI-GO-APTES composite was synthesized by wet chemical method, and its formation was confirmed using characterization techniques such as X-ray diffraction spectroscopy, scanning electron microscopy, X-ray photoelectron spectroscopy, thermal gravimetry analysis, electrochemical impedance spectroscopy and cyclic voltammetry. The characterization studies revealed that the synthesized PANI-GO-APTES had an amorphous structure, good electrical conductivity and good thermal stability. Due to its excellent electrochemical property, PANI-GO-APTES was investigated for electrochemical sensing application. PANI-GO-APTES cast on the glassy carbon electrode responded to Pb


Food choices are a key determinant of dietary intake, with brain regions, such as the mesolimbic and prefrontal cortex maturing at differential rates into adulthood. More needs to be understood about developmental changes in healthy and unhealthy food perceptions and preference. We investigated how food perceptions and preference vary as a function of age and how food attributes  impact age-related changes. One hundred thirty-nine participants  completed computerized tasks to rate high-calorie and low-calorie food cues for taste, health, and liking (preference), followed by 100 binary food choices based on each participant’s ratings. Dietary self-control was considered successful when the healthier (vs. tastier) food was chosen. Self-control success ratio was the proportion of success trials over total number of choices. Beta-weights for health  and taste (β-taste) were calculated as each attribute’s influence on food preference. Adiposity measurements included BMI z-score and waist-to-height ratio (WHtR). High-calorie foods were rated more tasty and less healthy with increasing age. Older participants liked high-calorie foods more (vs. younger participants), and β-taste was associated with age. Significant age-by-WHtR interactions were observed for health and taste ratings of high-calorie foods, β-taste, and marginally for preference of high-calorie foods. Stratifying by WHtR , we found age-related increases in taste and preference ratings of high-calorie foods in the high WHtR group alone. In contrast, age-related decreases in health ratings of high-calorie foods were significant in the low WHtR group alone. Age and β-taste were significantly associated in the high WHtR group and only marginally significant with low WHtR. Although participants rated low-calorie foods as less tasty and less healthy with increasing age, there was no association between age and preference for low-calorie foods. Participants made faster food choices with increasing age regardless of WHtR, with a significant age-by-WHtR interaction on reaction time (RT). There were no age-related effects in self-control success ratio and β-health. These results suggest that individual differences in age and central adiposity play an important role in preference for high-calorie foods, and a higher importance of food tastiness in food choice may contribute to greater preference for high-calorie foods with increasing age. Every day, we make a series of dietary decisions that determine energy intake . Over tiAge-related effects on the central regulation of eating have been shown in studies where younger individuals exhibit greater food craving for unhealthy foods compared to older individuals, decreased brain signal change in the prefrontal regions, and fewer connections between prefrontal-limbic regions implicated in regulation of eating , 2019. CIn addition to the imbalanced development of the prefrontal cortex and limbic system, adolescents could also develop less healthy eating behaviors with increasing independence . AdolescThe observed connections between brain development and dietary intake during adolescence present an opportunity for behavioral research in food decision-making. Dietary decision-making involves integration of basic food attributes, such as tastiness, and abstract attributes, such as healthiness. Food tastiness is reliably weighted in decisions, and is a primary driver of food choices in youth 8–14 years old, as studied with a computerized food ratings and subsequent food choice  task ; subjecta priori research question. At the time of initiating this study and its design in 2015, there were not any similar food choices studies in youth available in the literature to utilize as a basis for power calculation. Thus, sample size was largely driven by feasibility and resources. Our data collection ended due to the Covid-19 pandemic.This was a cross-sectional study of 139 individuals between 8 and 23 years old . Race was reported as 79.9% Caucasian, 11.6% Asian, 2.3% African-American, and 6.2% more than one race. For ethnicity 57.4% (74/129) were Hispanic or Latino. A subset of participants had both brain structural data and food choices behavioral data, for which we published their Self-Control Success Ratio variable data . Data weIn this study we included participants from age 8 to 23 years old. We included children starting at 8 years so that they would be able to understand and follow the instructions to complete the computer food ratings and food choice task, which were written at a third-grade level. Subjects 19–23 years old were included given recent proposals that the definition of adolescence be expanded to 24 years old to reflect continued neurodevelopment into the third decade of life , and giv2) and BMI z-score (BMI-z) was determined based on the U.S Center for Disease Control normative data1. Participants aged 20 and older had BMI-z calculated for 20 years of age. Waist circumference (n = 136) was measured at the midpoint between the iliac crest and lower costal margin in the midaxillary line, and waist-to-height ratio (WHtR) was calculated. Average BMI-z (SE) was 0.93 ± 0.09 and WHtR was 0.52 ± 0.01.Parents or subjects older than 18 years old filled out a demographic questionnaire. The height and weight of all participants were measured using a stadiometer and a calibrated digital scale, respectively. Body mass index (BMI) was calculated  for each food attribute on a 5-point verbal and visual Likert scale. Participants rated food tastiness as (1) really not tasty, (2) not tasty, (3) so-so, (4) kind of tasty, and (5) very tasty. Participants rated food healthiness as (1) really unhealthy, (2) not healthy, (3) so-so, (4) kind of healthy, and (5) really healthy. Participants rated preference as (1) really don’t like, (2) don’t like, (3) so-so, (4) kind of like, and (5) really like.Participants were shown 30 high-calorie and 30 low-calorie food cues and were asked to rate the cues according to tastiness (“How tasty is this food?”), healthiness , and preference or liking (“How much would you like to eat this food?”) . Block aBased on participants’ rating of the 60 food cues, a linear regression model was created for each participant to measure how well the health rating or taste rating predicted their preference rating for food cues. A beta weight for health  and for taste (β-taste) were determined for each participant from the 60 trials to quantify the relative influence given to each attribute in determining preference for a food cue.The ratings were then used to construct 100 binary pairs of food cues for the participant to choose between in the food-choice mouse-tracking task . Of thesEach participant’s reaction time, the time between the presentation of the food cues and when they indicated their choice, was also calculated. A self-control success ratio, the proportion of challenge trials in which the subject chose the healthier food cue over the tastier one, was calculated for each participant.t = 1 to t = 101) at which a one-sided t-test first reported the coefficient of the particular attribute in a multiple linear regression as significantly greater than 0. The individual Significance Times were corrected by first normalizing the particular multiple linear regression coefficient of the attribute to be a proportion of its final value, and then fitting these normalized coefficients to a logistic regression; the time window at which that regression first has a non-zero value is the corrected individual Significance Time.For each subject’s mean trajectory, a multiple linear regression was constructed to predict how differences in taste ratings and health ratings between the two food cues influenced the angle of the mouse from the start point. These models were then used to find the Significance Time for health and taste per participant, defined as the time at which the respective attribute  emerged and remained as a significant predictor of the participant’s final choice. The Significance Time was reported as the normalized time window , the furthest point of the mouse’s actual trajectory from the ideal trajectory , and the AUC, the space between the actual and ideal trajectories of the mouse’s path. MD represents how close a participant was to making a different decision while AUC is a measure of the cognitive effort with which the participants made decisions. The greater the AUC and MD, the greater the cognitive effort engaged to make a decision .Statistical analyses were conducted using SPSS . Linear regressions were used to examine relationships between age and food ratings. Food ratings included taste, health and preference of high-calorie and low-calorie food cues. We additionally examined relationships between age and β-taste and β-health using linear regressions.Mouse-tracking related variables included AUC for successful self-control trials, AUC for failed self-control trials, MD for successful self-control trials, and MD for failed self-control trials. As well, Significance Time for taste and for health. Relationships between age and self-control success ratio were investigated using linear regressions. We used linear regressions to investigate relationships between age and mouse-tracking related variables. We also examined age-related effects on reaction time during food choice using linear regressions.2) and linear models (age). We found that F change score was not significant for all the comparisons between quadratic models and linear models, suggesting that linear models fitted data better than quadratic models in age-related effects on food ratings and food choice data.Sex and WHtR were included as covariates in all the age-related models. For all the age-related models, age variable was transformed using a square-root transformation given the non-uniform distribution of age; we also explored if the quadratic model would fit the data better than the linear model of age-related effects on food ratings and food-choice mouse-tracking data. F change scores were compared between quadratic models  = 16.35, p < 0.001, partial eta squared = 0.11], such that low-calorie foods were rated higher than high-calorie foods . There was no significant main effect of attribute . There was a significant interaction of calorie and attribute . Post-hoc paired t-tests revealed that participants rated low-calorie foods more tasty  and healthier  than high-calorie foods. One-way ANOVA on food preference controlled for age, sex and WHtR revealed a significant main effect of calorie , such that participants liked low-calorie foods more than high-calorie foods .Across participants, the average rating ± SE of high-calorie foods in terms of tastiness was 3.52 ± 0.07, healthiness was 1.82 ± 0.04, and overall preference  was 3.40 ± 0.07. The average rating of low-calorie foods in terms of tastiness was 3.73 ± 0.06, healthiness was 4.42 ± 0.04 and overall liking was 3.70 ± 0.06. Repeated measures of ANOVA on food ratings with two-within subject factors  controlled for age, sex and WHtR revealed a significant main effect of calorie [p = 0.001). AUC and MD for successful self-control trials were larger than failed self-control trials  suggesting that greater cognitive effort was engaged during successful self-control choices than failed self-control choices.The average RT for food choices made was 1.63 ± 0.03 s. The average self-control success ratio for all participants was 33.55 ± 2.11%. Consistent with prior studies, we found that reaction time was longer during successful self-control trials than failed self-control trials , suggesting that the taste attribute is processed earlier than health attribute in food choice.Across participants, the average Significance Time for taste was 899.20 ± 34.50 milliseconds (ms), and the average Significance Time for health was 1043.59 ± 33.11 ms. We also found that the Significance Times for taste were earlier than Significance Times for health  and less healthy  with increasing age. As well, older participants indicated greater overall preference for high-calorie food cues  than the younger participants.High-calorie foods were rated as more tasty  and less healthy  with increasing age, there was no significant association between age and preference for low-calorie food cues .Although participants rated low-calorie foods as less tasty , suggesting that the taste attribute may contribute to the age-related increases in preference for high-calorie foods. There was no significant relationship between age and β-health .Older age was associated with an increased influence of taste attribute  on food preference . There was no significant association between age and self-control success ratio , neither were there significant associations between age and mouse-tracking related variables .Participants made faster food choices with increasing age  and health  ratings for high-calorie foods, and a marginally significant interaction for preference ratings  for high-calorie foods  and preference  for high-calorie food cues were significant in the high WHtR group, but not in the low WHtR group . In contrast, age-related decreases in health ratings of high-calorie foods were significant in the low WHtR group , but not in the high WHtR group .There was a significant interaction of age-by-WHtR on taste . When data were stratified into high and low WHtR groups a significant association between age and β-taste was observed in the high WHtR group , and a marginally significant relationship between age and β-taste was found in the low WHtR group   .SE = 0.11, p = 0.21, partial eta squared = 0.01), health  and preference ratings  for low-calorie foods as well as β-health .We did not find significant age-by-WHtR interactions on taste , such that both the high  and low  WHtR groups exhibited a faster reaction time with increasing age. There was a significant age-by-WHtR interaction on AUC . When data were stratified into high and low WHtR groups, we found no significant relationships between age and AUC in both high  and low WHtR  groups.A significant age-by-WHtR interaction was observed on reaction time , MD , Significance Time for taste , and Significance Time for health .We did not observe significant age-by-WHtR interaction on self-control success ratio  could be due to lack of power. As well, positive results need to be interpreted with caution given there might be a possibility of type 1 error induced false positive results. There are also inherent limitations in all laboratory food choices research, as they may differ from real-life choices. Finally, future correlation of behavioral task results with brain imaging data would be useful in further understanding neurobiological underpinnings of developmental trajectory of food decision-making.We conclude that our results are consistent with other studies that demonstrate age-related increases in consumption of calorie-dense foods in youth, in particular in those with central obesity, and suggest that age and adiposity may be more relevant to preference for high-calorie foods. Interventions targeting youth at an early age could therefore be beneficial to helping reduce consumption of high-calorie foods over time.The raw data supporting the conclusions of this article will be made available by the authors, upon reasonable request.The studies involving human participants were reviewed and approved by the Institutional Review Board of CHLA and USC (CHLA-15-00007 and HS-16-00978). Written informed consent to participate in this study was provided by the participants’ legal guardian/next of kin.MSG, MK, and SLu took responsibility for the integrity of the data in the study and the accuracy of the data analysis, and drafted the manuscript. SLu contributed to statistical analysis. MK, MH, and SLu provided administrative, technical, or material support, supervised the work, and obtained funding. All authors contributed to acquisition, analysis, or interpretation of data and critical revision of the manuscript for important intellectual content.The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH, USC DORI, or the Stewart Clifton Endowment.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


To assess if quantitative integrated deep learning and radiomics features can predict the PD-L1 expression level in preoperative MRI of hepatocellular carcinoma (HCC) patients. The data in this study consist of 103 hepatocellular carcinoma patients who received immunotherapy in a single center. These patients were divided into a high PD-L1 expression group (30 patients) and a low PD-L1 expression group (73 patients). Both radiomics and deep learning features were extracted from their MRI sequence of T2-WI, which were merged into an integrative feature space for machine learning for the prediction of PD-L1 expression. The five-fold cross-validation was adopted to validate the performance of the model, while the AUC was used to assess the predictive ability of the model. Based on the five-fold cross-validation, the integrated model achieved the best prediction performance, with an AUC score of 0.897 ± 0.084, followed by the deep learning-based model with an AUC of 0.852 ± 0.043 then the radiomics-based model with AUC of 0.794 ± 0.035. The feature set integrating radiomics and deep learning features is more effective in predicting PD-L1 expression level than only one feature type. The integrated model can achieve fast and accurate prediction of PD-L1 expression status in preoperative MRI of HCC patients. Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related death worldwide . It is tRadiomics, a high-dimensional quantitative feature analysis approach, can extract high-throughput features from medical imaging and perform quantitative analysis of tumor heterogeneity. Several studies have shown that radiomics features could characterize the tumor and its tumor microenvironment (TME) ,40,41,42In this study, we propose an integrated model for assessing PD-L1 expression level in HCC patients by integrating deep learning and radiomics features in preoperative MRI, and then carry out a quantitative analysis of the features. We only use the deep learning model as a feature extractor (not a classifier), and then integrate the deep learning features into the radiomics analysis model, which enriches the predictive power of the model and improves the overall performance of the model with limited training data. Our goal is to assess whether quantitative integrated deep learning and radiomics features can predict the PD-L1 expression level in preoperative MRI of HCC patients.This is a retrospective single-center study, that consisted of 103 HCC patients who received treatment in our center between December 2017 and May 2021. The hospital’s ethical review committee approved the study. The inclusion criteria of cases enrolled were as follows: (1) patients with pathologically proven HCC; (2) abdominal MRI performed within one month before surgery; (3) availability of tumor samples and clinical data; (4) IHC examination of PD-L1 performed. Exclusion criteria were (1) the interval between MRI examination and surgery was more than 1 month; (2) poor MR imaging quality; (3) patients with other tumor diseases than HCC at the same time.The PD-L1 expression level was measured by the IHC test with a detection kit of Ventana PD-L1 (SP142). The test specimen was paraffin tumor tissue obtained by surgical intervention. The specimen was fixed by a concentration of 10% neutral formalin for 6–8 h, regarding full circumferential or partial cell membrane staining as PD-L1 positive tumor cells. The PD-L1 high expression was defined as the percentage of PD-L1-positive tumor cells relative to the total tumor cells exceeding 50%; otherwise, it was defined as PD-L1 low expression. Finally, the 103 enrolled patients were divided into two groups by PD-L1 expression level: a high PD-L1 expression group (30 patients) and a low PD-L1 expression group (73 patients).All MR images were acquired on a 3.0T MRI scanner device . The MRI protocol consisted of T2 weighted TSE sequence . The segmentation of the entire tumor volume of interests (VOI) was manually performed slice by slice on T2-WI by a radiologist with 10 years of experience; after that, each segmentation slice was reviewed and modified by a chief radiologist, who had over 20 years of experience in MRI. The region of interest (ROI) covered the whole tumor. The Medical Imaging Interaction Toolkit (MITK) v2018.04 software was applied to draw the tumor VOI. A total of 1595 3-D radiological features were extracted from each VOI with Pyradiomics . These r−3 and 50 epochs.A total number of 1024 deep learning features for each patient were extracted from a three-dimensional convolution neural network (3D-CNN), which consists of two 3D convolution layers and two fully connected layers. The deep features were obtained from the outputs of the first fully connected layer after applying the rectified linear unit (ReLU)  activatiThe output features of the first fully connected layer of the 3D-CNN framework were connected with the latter half of the radiomics process so that the extracted 1595 radiomics features and 1024 deep learning features were merged into an integrative feature space. The total number of features after fusion was 2619, as shown in We applied normalization to the feature matrix. Each feature vector was subtracted from the average value of the vector and then divided by its length. Due to the high dimensionality of the feature space, we utilized the Pearson correlation coefficient (PCC)  analysisp-value < 0.05 was considered significant and p-value < 0.01 was extremely significant. The ROC curve was used to assess the predictive ability of the model, and its corresponding quantification was calculated by the AUC. The accuracy, recall, specificity, precision, and f1_score were also calculated at the cutoff value that maximized the AUC value [The univariate analysis method was used to evaluate the statistical significance of patients’ clinical characteristics in the prediction of PD-L1 expression. A two-sided UC value . To valip > 0.05 for all)  with tumor cells expressing PD-L1 ≥ 50% were assigned into the high expression group, and 73 patients (age median: 54.48 ± 12.70) with expression scored < 50% were assigned to the low expression group. In our univariate analysis, there was no significant difference in clinical characteristics including Age, Sex, HBV_DNA, HBs, AFP, Maximal tumor diameter, CEA, and TBIL between the two PD-L1 expression groups  .The experiment compared the classification performance of different classifiers, and the best-performing classifier was used to further verify the performance of strategies based on different features using five-fold cross-validation. The compared classifiers are SVM, AE, LR-Lass, Decision Tree, Random Forest, and LDA. All classifiers were trained  and tested  on the same dataset. The predictive results are shown in The integrated predictive model with the best AUC is based on 14 selected features, and the details are shown in RsF+DLF_Score = , deep_feature_841 (p = 0.006), deep_feature_193 (p = 0.012), deep_feature_670 (p = 0.019), deep_feature_81 (p = 0.028) and deep_feature_805 (p = 0.031). The results of statistical significance indicate that these characteristics are correlated with the expression level of PD-L1. The number in the deep_features represents the order of neurons in the fully connected layers. For example, deep_feature_524 indicates that this feature is taken from the 524th neuron of the fully connected layer. As shown in the column of coefficients in p-value < 0.01); its scores in the high expression group are mainly distributed in the positive value area, while the group in the low expression group are distributed in the negative value area, and there is a significant difference in distribution between the two groups. Among the features of radiomics, original_glcm_InverseVariance and wavelet_HHL_glcm_InverseVariance are strongly related to PD-L1 expression prediction with p values of 0.001 and 0.001, respectively. In addition, their coefficients in SVM linear function are all negative, with value of −0.995 and −0.748 (in p = 2.502 × 10−9. The score is higher in the high expression group (0.27 ± 0.44) than in the low expression group (−0.11 ± 0.33) as shown in Several MRI radiomics features and deep learning features had shown correlation with PD-L1 expression. In deep learning features, six deep learning features have statistical significance to expression level of PD-L1, which are deep_feature_524 (0.748 in . It indiHowever, there are some limitations to our study. First, it is a retrospective study from a single center without additional validation by other hospitals, and the number of HCC cases with the PD-L1 test was limited, especially the proportion of patients with high PD-L1 expression. Therefore, the validity of our research results might have been impaired. In future research, more cases and multi-center datasets are needed. Second, although we have given the features that contribute to predicting PD-L1 expression status in the combined model, only the features of radiomics are interpretable, while the interpretability of the deep learning features is still poor. The possible correlation between the tumor biological mechanisms and the deep learning features will be investigated from molecular, proteins, and gene levels in future work. Additionally, due to the data collection issue, only T2-WI sequence of MRI was available to be analyzed in this study, which may neglect information from other kinds of sequences.We proposed a prediction model of integrating the features of radiomics and deep learning for quick and accurate assessment of PD-L1 expression levels for HCC patients before ICI therapy. Our integrated model achieves good performance on predicting PD-L1 expression level in preoperative MRI. These results suggest that MRI radiomics combined with deep learning analysis shows potential for clinical practice in the assessment of PD-L1 expression in HCC, which may achieve the stratification of HCC patients for immunotherapy.


Calcaneal apophysitis pain perception was improved in both groups, but children who used custom-made foot orthoses showed a greater improvement.The aim of the present research was to determine the effectiveness of relieving calcaneal apophysitis pain using “off-the-shelf” heel-lifts and custom-made orthotics. Two intervention modalities were evaluated and compared in a 12-week follow-up trial. Inclusion criteria included 9- to 12-year-old children diagnosed with calcaneal apophysitis. Children were randomly stratified into treatment A (custom-made polypropylene foot orthoses) and treatment B (“off-the-shelf” heel-lifts) groups. Treatment effectiveness was measured by algometry and the visual analogical scale (VAS). A total of 208 patients were included. The treatment A group showed an increase in threshold algometry of 53.4% (95% CI 47.1% to 59.7%) and a decrease in VAS of −68.6% (95% CI −74.5% to −62.7%) compared with the treatment B group ( Calcaneal apophysitis (Sever´s disease) is a very common ailment present in the heels of children between 7 and 15 years old ,3,4,5. TCalcaneal apophysitis, fortunately, is a benign ailment which disappears without exception after puberty when the secondary ossification center of the calcaneus is closed . UnfortuThe described conservative strategies are popular  but presAlong this line, there are some studies in the literature that have used orthoses to act both on ACPS and repetitive impacts ,14,15,162), lunge test , foot posture index (FPI-6) , and visual analogical scale (VAS)score  as these are considered to be important risk factors associated with pain severity in calcaneal apophysitis [The study was a parallel-group, randomized comparative effectiveness trial with concealed allocation, blinding of investigators and assessors, and intention-to-treat analysis. It examined the effect of custom-made foot orthoses and heel-lifts in children with calcaneal apophysitis. Participants were enrolled at the time of calcaneal apophysitis diagnosis. After baseline assessment, children were individually randomized to “custom-made foot orthoses” (treatment A group) or “off-the-shelf heel-lifts” (treatment B group). Concealed allocation was carried out by having randomization performed by a third party who was not involved in the recruitment or treatment of the children. Eight permuted blocks were used to stratify randomization by body mass index (BMI)  consisted of a treatment period of 12 weeks, following the design of previous studies . Both inChildren in the treatment A group received custom-made foot orthoses as treatment intervention. The orthoses had a thickness of 9 mm and were composed of the following materials: confortene, polypropylene, poron XRD, and lunasoft . These kChildren in the treatment B group received an 8 mm “off-the-shelf” heel-lift as treatment intervention. These were composed of confortene, poron XRD, and ethyl vinyl acetate (EVA) . ChildreChildren were blinded to group allocation. Both treatment A and B groups received the same attention during the consultation, and they were informed that the intervention given was adequate for their disease before treatment application and after the follow up.The primary outcome and the characteristics of the sample of the present research were obtained at baseline and after 12 weeks of follow up by an experienced and trained assessor who was blinded to group allocation. At baseline, participants’ weight and height were measured with an Año-Sayol scale and stadiometer, respectively .®, Greenwich, CT 06836-1217 USA) was used. It has been used before in children with calcaneal apophysitis [The primary outcome was calcaneal apophysitis pain perception, which was determined by the VAS and the algometry threshold. The VAS is commonly used to measure pain perception  and has physitis . To perfphysitis .Values for the BMI, FPI-6, and lunge test in the children with calcaneal apophysitis were taken as the baseline characteristics of the sample. The FPI-6 was performed following the guidelines of Redmond et al. . FPI is Sample size estimation was based on the detection of a difference of 29% in the number of children with pain and a mean difference of 0.39 or higher for the algometry and the VAS between baseline and after the follow up based on a test of a hypothesis comparing proportions between two related groups. Assuming a two-sided α of 0.05, power of 80%, and a 10% drop-out rate, a sample size of 198 participants was required. Continuous variables were expressed as the mean ± standard deviation (SD), whereas qualitative variables were expressed as frequencies and percentages. Continuous data were checked for normality by the Kolmogorov–Smirnov test.The baseline characteristics of the children were summarized using descriptive statistics and tabulated for comparison between both groups. Baseline characteristics were compared between included and excluded participants to analyze whether they were representative. Mann–Whitney U and Student’s t-tests were used to compare continuous variables, and the Fisher Test was used to compare dichotomous variables.Differences between baseline and final assessment in algometry and VAS were performed with the calculation of the “percentage change”. Positive results indicated an increase in values at the end of the study, and negative results indicated a decrease. Treatment effectiveness after the follow up was determined by regression models. Linear regression was applied to quantitative variables, algometry, and VAS, and logistic regression was applied to dichotomous qualitative variables; association was measured in terms of odds ratio.p-value < 0.05 was considered significant. The statistical analyses were performed using the SPSS software 22.0 for Windows .Statistical analysis of the outcomes was performed according to intention-to-treat principles, comparing the groups regarding pain perception evolution. For all tests, a two-sided Between June 2019 and February 2020, 234 children were screened, 208 of which were included in the present study. Among these, 104 children were allocated to both treatment A and B groups. The flow of participants throughout the study is presented in Children had a reasonable compliance with their allocated interventions. Only four and five children were lost to follow up in the TA and TB groups, respectively . These cp < 0.05) for both interventions. The values for VAS decreased by 68.5 ± 15.4 points in the treatment A group (p < 0.001) and 14 ± 17.7 points in the treatment B group (p < 0.001); the algometry values increased by 2.0 ± 0.5 kgf in the treatment A group (p < 0.001) and 0.6 ± 0.6 kgf in the treatment B group (p < 0.001). Differences in changes and improvement between treatment A and B groups are shown in p < 0.001). Results were compared between groups using odds ratio, and the confidence intervals were wide, as shown in The primary outcome of calcaneal apophysitis pain perception analysis is shown in The odds ratio (95% CI) between groups showed that children who wore custom-made foot orthoses had a higher improvement, which increased algometry data by 53.4 (47.1 to 59.7) and reduced VAS by 68.6 (74.5 to 62.7), compared with children who wore heel-lifts.The purpose of this trial was to compare heel pain perception in children with calcaneal apophysitis using custom-made polypropylene foot orthoses and “off-the-shelf” heel-lifts in an intervention period of 12 weeks. Calcaneal apophysitis pain perception for the three variables measured by VA and algometry were significantly improved and reduced in both groups. The treatment A group showed significant pain relief compared with the treatment B group.At baseline, all the participants had high VAS values and a reduced pressure pain threshold on the affected heel. Pain relief was significantly different between treatment A (custom-made foot orthoses) and treatment B (heel-lifts) groups.The heel-lift’s function was to lift the heel with an inclined plane, which allowed a reduction in Achilles tendon tension and traction on the calcaneus bony surface ,8,9,10. p < 0.001). Compared with the treatment B group, the treatment A group experienced an increase in the algometry threshold of 53.4% and a VAS punctuation reduction of −68.6%. Similar results were obtained in 2011 in two studies performed by Perhamre et al. [Improvement in the treatment B group was found in approximately 20–30% of children, while in the treatment A group, it was found in 70–90% of children  were homogeneous, and the study provided consistent data about the use of custom-made foot orthoses for calcaneal apophysitis pain relief.The potential use of custom-made foot orthoses in the approach to calcaneal apophysitis needs to be considered in clinical consultation as an effective treatment strategy. Future studies should consider the comparison between custom-made foot orthoses against other treatment strategies and protocols, such as strengthening and stretching exercises, physiotherapy approaches, drug treatment, and physical exercise tests, for the evaluation of pain perception.In summary, this study highlights that the use of custom-made foot orthoses instead of heel-lifts for calcaneal apophysitis (Sever´s disease), applied during a 12-week follow-up period, may have a substantial effect on calcaneal apophysitis pain relief.


The year 2013 marks a watershed in the history of medical education in Taiwan. Following Taiwan’s Taskforce of Medical School Curriculum Reform recommendations, the medical school curriculum was reduced from 7 to 6 years. This study aimed to analyze the impact of medical school curriculum reform on medical students’ performance in objective structured clinical examinations (OSCEs).We retrospectively analyzed the OSCE records at Taipei Veterans General Hospital (Taipei VGH), one of Taiwan’s largest tertiary medical centers, between November 2016 and July 2020. The eligibility criteria were medical students receiving a full one-year clinical sub-internship training at Taipei VGH and in their last year of medical school. All medical students received a mock OSCE-1 at the beginning of their sub-internship, a mock OSCE-2 after six months of training, and a national OSCE at the end of their sub-internship. The parameters for performance in OSCEs included “percentage of scores above the qualification standard” and “percentage of qualified stations.”p < 0.001), and mock OSCE-2 . Moreover, medical students in the 7-year curriculum had a higher percentage of qualified stations in mock OSCE-1  and mock OSCE-2 . After clinical sub-internship training, there were no differences in the percentage of scores above the qualification standard  and percentage of qualified stations .Between November 2016 and July 2020, 361 undergraduates underwent clinical sub-internship training at Taipei VGH. Among them, 218 were taught under the 7-year curriculum, and 143 were instructed under the 6-year curriculum. Based on baseline-adjusted ANCOVA results, medical students under the 7-year curriculum had a higher percentage of scores above the qualification standard than those under the 6-year curriculum at the mock OSCE-1 (7-year curriculum vs. 6-year curriculum: 33.8% [95% CI 32.0–35.7] vs. 28.2% [95% CI 25.9–30.4], At the beginning of the sub-internship, medical students under the 7-year curriculum had better OSCE performance than those under the 6-year curriculum. After the clinical sub-internship training in Taipei VGH, there was no difference in the national OSCE score between the 6- and 7-year curricula. Our study suggests that clinical sub-internship is crucial for the development of clinical skills and performance in the national OSCE. Curricular reform in medical schools has become a popular topic in recent decades –3. In TaShortening the medical curricula is an international phenomenon. For example, in the United States, there have been three waves of shortening the curricular length of medical schools. In the United States, a 3-year curriculum was introduced in the 1940s and 1970s . In the We retrospectively analyzed the data of mock and national OSCEs of undergraduates at Taipei Veterans General Hospital (VGH), a tertiary medical center in Taiwan, between November 2016 and July 2020. All undergraduates received two mock OSCEs and one national OSCE during their sub-internships. The mock OSCE-1 and mock OSCE-2 are given two months and six months after the beginning of the sub-internship, respectively. Finally, students take the national OSCE in the last month of training.The procedure of OSCEs has been validated and widely applied in Taiwan’s teaching hospital for more than ten years . All exaIn this study, we used two outcome measurements for medical students’ OSCE performance. The percentage of scores above the qualification standard is defined as the difference between the students’ actual score and the approval standard score, divided by the approval standard score. In Taiwan, the determination of the approval standard score is based on the examinee-centered borderline group method with regression, following the standard protocol of Taiwan’s national OSCE board , 18–23. p < 0.05. All statistical analyses were conducted using IBM SPSS (version 22.0).Relationships were analyzed between outcome measurements and different curricula, as well as demographics. Means were reported with standard deviations, and medians were reported with interquartile ranges. Comparisons of performance in serial OSCE were analyzed using repeated-measures ANOVA. To compare the OSCE performance between the two curricula, a two-way analysis of covariance (ANCOVA) for repeated measures was performed with the baseline scores used as a covariate to eliminate the possible demographic influence of OSCE performance . Relatiop < 0.001) (Table p < 0.001), and the percentage of qualified stations   underwent clinical internship training at Taipei VGH. Among them, 218 were under the 7-year curriculum, and 143 were under the 6-year curriculum. Comparing the two curriculum lengths, students under the 6-year curriculum had a higher percentage of clerkship training at our institution  undergraduate students received clerkship training at our institution. Among baseline demographics, history of received clerkship at our institution was associated with better performance in OSCEs in two outcome measurements Table .Table 3Ap < 0.001), and a higher percentage of qualified stations . In the mock OSCE-2 (after six months of training), medical students under the 7-year curriculum had a higher percentage of scores above the qualification standard than those under the 6-year curriculum , and a higher percentage of qualified stations . In the national OSCE, there were no differences in the percentage of scores above the qualification standard  and the percentage of qualified stations   than those under the 6-year curriculum (7-year curriculum vs. 6-year curriculum: 33.8% 95% CI 32.0–35.7] vs. 28.2% [95% CI 25.9–30.4], .0–35.7 vIn this study, we found medical students under the 7-year curriculum performed better in OSCE than their 6-year curriculum counterparts at the beginning of the internship. After the clinical internship training at Taipei VGH, there was no difference in national OSCE scores between the 6- and 7-year curricula graduates.There is a paucity of controlled studies comparing the outcomes between different medical school curriculum lengths . TherefoThe strength of the present study is that we used standardized OSCEs as outcome measurements. This OSCE protocol has been integrated into the national medical examination in Taiwan, which is a more objective measurement for the educational outcome . AnotherOne critical issue that should be addressed is the decline of the 2nd OSCE shown in Table At the beginning of the sub-internship, medical students under the 7-year curriculum outperformed those under the 6-year curriculum on the OSCE. After the sub-internship training, there was no difference in the national OSCE score between the 6- and 7-year curricula. Our study showed that changes in curriculum length in medical schools did not interfere with the OSCE performance at the end of the sub-internship, and clinical training is a crucial factor for developing clinical competencies. Our experience can inspire future curriculum reforms in medical schools.


Emulsion gel, a novel oral delivery carrier, provides the possibility to co-load hydrophilic and lipophilic nutrients simultaneously. In this study, duo-induction methods of laccase and glucono-δ-lactone (L&GDL) or laccase and transglutaminase (L&MTG) were used to prepare the soy protein isolate-sugar beet pectin (SPI-SBP) emulsion gel. The textural data of the emulsion gel was normalized to analyze the effect of different induction methods on the gel property of the SPI-SBP emulsion gels. The characterization studies showed the structure of L&MTG emulsion gel was denser with a lower swelling ratio and reduced degree of digestion, compared with L&GDL emulsion gel. Moreover, the release profiles of both β-carotene and riboflavin co-loaded in the SPI-SBP emulsion gels were correlated to the digestion patterns of the gel matrix; the controlled-release of encapsulated functional factors was regulated by a gel network induced by different induction methods, mainly due to the resulting porosity of the structure and swelling ratio during digestion. In conclusion, SPI-SBP emulsion gels have the capability of encapsulating multiple functional factors with different physicochemical properties. Designing a bio-degradable and safe delivery system for nutrient delivery has attracted much attention in recent years . TraditiFood-grade emulsion gels are usually constructed by biomacromolecules such as proteins and polysaccharides. Proteins are amphiphilic and are therefore often used as an emulsifier in an oil-water system. Although research on emulsion gels fabricated by whey protein isolate (WPI) , soy proProtein-polysaccharide emulsion gels can be fabricated through methods such as physical alterations, ionic strength, pH, and applications of enzymes . For lowThe current study aims to prepare a novel SPI-SBP emulsion gel as a multi-phased carrier to co-encapsulate both hydrophilic and lipophilic nutrients. The effect of different duo-induction methods using laccase in combination with GDL or MTG on the structural properties of SPI-SBP emulsion gel was evaluated. The digestive properties of the selected SPI-SBP emulsion gels for the subsequent release rate of bioactive substances were also measured. The establishment of a relationship among the gelation mechanisms, texture, microstructure and digestive profiles was attempted. The current study broadens the application of SPI-SBP emulsion gels as a multi-phased nutrient carrier to encapsulate thermal-sensitive substances.Soy protein isolate (SPI) was supplied by Shandong Yuwang Ecological Food Industry Co., Ltd. . Sugar beet pectin (SBP) was purchased from Herbstreith & Fox KG . Medium-chain triglycerides (MCT) was obtained from Guangzhou Yeshang trade Co., Ltd. . Laccase (≥0.5 U/mg) was provided by Sigma-Aldrich Co. Ltd. . Glucono-δ-lactone (GDL) was purchased from Beijing Solarbio Science & Technology Co., Ltd. . Transglutaminase (MTG) (≥1.5 U/mg) was supplied by Kuer Chemical Technology Co., Ltd. . All other chemicals were of analytical grade.w/w) was dissolved in an SPI solution  and the mixture was stirred for 4 h at room temperature. After being stored at 4 °C overnight, the SPI-SBP solution was heated at 85 °C for 15 min before being cooled down to room temperature immediately. Then, 20% MCT (w/w) was added to the SPI-SBP solution, and the mixture was mixed with a high-speed shear machine  at 10,000 r/min for 3 min, followed by homogenization five times at 30 MPa with a homogenizer  to obtain the SPI-SBP emulsion. The emulsion was mixed with GDL  and stirred for 1 min to yield the GDL SPI-SBP emulsion, or mixed with MTG (5–30 U/g protein) to obtain the MTG SPI-SBP emulsion. Next, a 20 U/g substrate of laccase was added into GDL or MTG SPI-SBP emulsion and stirred for 1 min to obtain L&GDL or L&MTG SPI-SBP emulsion. A control emulsion was prepared with 20 U/g substrate of laccase with SPI-SBP emulsion to illustrate the advantages of duo-induced emulsion gels in gel properties and functional factor delivery. Finally, the laccase, L&GDL, or L&MTG emulsion gels were prepared after heating the L&GDL or L&MTG emulsion at 40 °C for 4 h. All prepared gels were settled at 4 °C overnight for the subsequent experiments.SBP  and the 20 mm diameter P/20a cylindrical probe were performed to measure the textural properties of the SPI-SBP emulsion gel. The TPA parameters were as follows: the pressing height was 25% of the sample height, and the test speed was 60 mm/min. The TPA curves were recorded and the parameters  were calculated using the software provided by the Texture Analyzer .The original texture parameters  were normalized based on the Minkowski distance to obtain the comprehensive property index (CPI) of all samples . The corThe previously obtained SPI-SBP emulsion gel was freeze-dried before being observed on a scanning electron microscope  at a voltage of 15 kV.w/v) rhodamine B and 0.1% (w/v) Nile Red mixed staining solution was added into 2 g of the prepared SPI-SBP emulsion, after which the gelling agents were added according to the method described in Based on a previously published method , SBP was1 is the weight of the fresh emulsion gel and w2 is the weight of the emulsion gel after soaking.To assess the swelling property of the prepared emulsion gel, fresh prepared SPI-SBP emulsion gel was soaked in a 0.02% sodium azide solution  for 48 h , and thew/v) was targeted. After that, the same volume of simulated gastric fluid  was added to the oral mixture. Following this, 2000 U/mL of porcine pepsin (EC 3.4.23.1) was added to the mixture. The mixture was incubated at 37 °C for 120 min. After gastric digestion, the gastric chyme was mixed with the same volume of simulated intestinal fluid . The activity of trypsin (EC 3.4.21.4) and pancreatic lipase (EC 3.1.1.3) was 100 U/mL and 2000 U/mL in the final mixture, and the final concentration of bile salts was 10 mmol/L. The final mixture was incubated at 37 °C for 120 min. All incubating processes were performed in a 37 °C water bath shaker with a speed of 120 rpm [The SPI-SBP emulsion gels were cut into 0.2 cm × 0.2 cm × 0.2 cm cubes. These cubes were firstly incubated in simulated saliva fluid  at 37 °C for 5 min, and the ratio of gel cubes to SSF of 50:50 . Next, 10 μL of the supernatant was added into a 96-well microtiter plate, and 100 μL of ninhydrin reagent (20 mg/mL) dissolved in acetate buffer  was added into the mixture; the resulting microtiter plate was heated at 104 °C for 10 min. The absorbance was measured at 575 nm using a microplate reader , and glycine standards were used to calculate the amount of FAA in the samples.The determination of FAA was based on the method published by Rui . After iIn this study, a titration method reported by Sarkar  was usedw/w)) was dissolved in MCT at 45 ℃ to prepare the emulsion oil phase. Steps to prepare SPI-SBP emulsions and emulsion gels  co-loading systems were carried out following the steps described in Based on g for 10 min. The n-hexane phase was collected and the absorbance was measured at 450 nm using a microplate reader . The β-carotene standards were used to calculate the amount of β-carotene in the hexane layer [1 (mg) is the total amount of nutrients, w2 (mg) is the amount of nutrients in the supernatant.Freshly prepared emulsion samples and emulsion-gel samples loaded with nutrients were freeze-dried. A total of 0.5 g of each freeze-dried sample was soaked in 10 mL DI water for 24 h to release the embedded nutrients. Following this, 10 mL of n-hexane was added to the mixture, which was centrifuged at 8500× ne layer ,26. The 1 (mg) is the total amount of nutrients, w2 (mg) is the amount of nutrients in the supernatant.The co-loading systems of the SPI-SBP emulsion and emulsion gels  were digested based on the digestive procedure described in p < 0.05). Matlab 2016 software was used to analyze and normalize the textural data statistically and to perform a regression analysis as well as Pearson correlation analysis. The experimental data were processed using Origin 2019 software, and the results were expressed in means ± standard deviation. All experiments were conducted in triplicate.SPSS Statistics 26.0 software was used to analyze the data, and one-way analysis of variance (ANOVA) was used to determine the significant difference between each test (The textural properties were characterized by TPA, as shown in On the other hand, the hardness D, spring2 of the equation was 0.9785, which suggested that the linear regression model fitted the textural property data and could reflect the texture properties of the SPI-SBP emulsion gel.Based on the Minkowski distance, the three-dimensional data of the gel textural properties were normalized into one-dimensional data, which was defined as CPI of the SPI-SBP emulsion gel . AccordiAs shown in In A previous study indicated that the greater the swelling, the easier it is for water and digestive enzymes to diffuse into the gel network structure, which in turn promotes the release of functional factors . As showAs shown in After experiencing shear effects in the mouth and stomach, the oil in the emulsion is mixed with pancreatic lipase, trypsin, bile salts, and phospholipids in the small intestine. Oil digestion under the action of lipase was mainly carried out in the small intestine , so the EE is essential to the performance of the nutrient delivery system, and affects the storage and bioavailability of functional factors that are embedded in the delivery system . In thisThe release of β-carotene in the SPI-SBP emulsion gel during the digestion process is mainly related to the digestion process of oil embedded in the gel network . After sAs indicated in In this study, a novel SPI-SBP emulsion gel was prepared as a multi-phased carrier to co-encapsulate both hydrophilic and lipophilic nutrients. Compared with the L&GDL emulsion gel, the structure of the L&MTG emulsion gel was denser and had a lower swelling ratio; therefore, the digestion rate was lower. The EE of β-carotene in the SPI-SBP emulsion gel was not affected by the induction method, while the EE of riboflavin was mainly correlated to the pore size of the SPI-SBP network synthesized under different induction methods. During simulated digestion studies, the release of both β-carotene and riboflavin in the SPI-SBP emulsion gels was regulated by the gel network induced by different induction methods. The swelling ratio accompanied by these structures was responsible for the controlled-release profiles of functional factors co-loaded within the system. Therefore, the capability of SPI-SBP emulsion gels of encapsulating multiple functional factors with different physicochemical properties will further expand their applications in the food industry.